Field of the Invention (Technical Field)
The present invention relates to a method of measuring cancer-related substances which increases in blood with the progress of cancer, including mainly a free DNA (DNA wrapped around the histones) as a target to be measured by Raman spectroscopy.
Description of Related Art
A method of measuring a cancer-related substance which increases in the blood with the progress of the disease has hitherto been used as a diagnostic method for cancer. The cancer-related substances referred to proteins and the like, which are cancer-specific substances extracted from a body fluid of cancer patients and are generally released into the blood when the cancer cells are destroyed. According to the prior diagnostic methods of the cancer, it is determined that there is a possibility that the test subject or patient is suffering from cancer when more than a determined value of the cancer-related substance is found existing in the blood.
Thus as the cancer-related substance released into the blood by destruction of the cancer cells, it is known that not only proteins but also DNA may be released into the blood. And, when compared with the healthy subjects and cancer patients, it has been reported that the amount of the free DNA (ctDNA) derived from cancer cells in blood, is significantly more in those cancer patients than healthy individuals. Thus, by quantifying the free DNA of cancer cells from the body fluid such as blood, it is considered to be able to diagnose the presence of cancer. As such a method of cancer diagnosis, for example, there are proposed 1) a method of diagnosing a possibility of cancer in case of detecting 200 bp or more of DNA to be amplified by the polymerase chain reaction (PCR) method and the like, in the body fluid or feces discharged from the body, and further analyzing a mutation in its DNA if necessary (Patent Document 1 and 2), and 2) a method of quantifying genomic DNA contained in a body fluid, and further performing DNA testing in the case of more than a predetermined value of the genomic DNA (Patent Document 3).
Furthermore, even if the patient is diagnosed suffering from cancer, mere quantitative analysis of the DNA in body fluids is unable to identify a cancer suffering organ. When the cancer is arising and progressing, it is known that a specific mutation of DNA occurs depending on the original cancer site. Therefore, by clarifying the type of mutation in the DNA, it may be possible to identify an organ or a cancer site where the cancer is developing. Here, as Mutations of DNA, there are listed point mutations of DNA and also structural abnormalities such as chromosome gain or loss. For example, in about 70% of pancreatic cancer, it is known that the point mutation occurs in the K-ras gene. Also, in the analysis of loss of heterozygous, (hereinafter referred to as LOH) there have been reported the loss of specific chromosomal arms depending on each cancer type, for example, it is known that LOH is concentrated on the short arm of chromosome 3 in case of the lung cancer. Also, the amplification of long arm No. 8 of chromosome and the amplification of RB2 are known in the breast cancer, Therefore, in order to provide an improved method for diagnosing cancer with high accuracy by quantifying the free DNA from cancer cells with a biochip as described herein, there has been provided a method of diagnosing cancer, which comprises a step of extracting a free DNA from plasma collected from a subject, a step of calculating the free DNA per unit volume of the extracted plasma by quantifying the free DNA, a step of comparing the calculated value of the free DNA with a second threshold value more than the first threshold value, a step of making a diagnose as follows; the subject has a high possibility of affection with cancer when the calculated value is less than the first threshold value, while some DNA from normal cells are mixed in the plasma when the above threshold is more than the second threshold value (patent document 4).